Search

Genetic variants that are associated with susceptibility to type 2 diabetes (T2D) are important for identification of individuals at risk and can provide insights into the molecular basis of disease. Analysis of T2D in domestic animals provides both the opportunity to improve veterinary management and breeding programs as well as to identify novel T2D risk genes. Australian-bred Burmese (ABB) cats have a 4-fold increased incidence of type 2 diabetes (T2D) compared to Burmese cats bred in the United States. This is likely attributable to a genetic founder effect. We investigated this by performing a genome-wide association scan on ABB cats. Four SNPs were associated with the ABB T2D phenotype with p values &lt, 0.005. All exons and splice junctions of candidate genes near significant single-nucleotide polymorphisms (SNPs) were sequenced, including the genes DGKG, IFG2BP2, SLC8A1, E2F6, ETV5, TRA2B and LIPH. Six candidate polymorphisms were followed up in a larger cohort of ABB cats with or without T2D and also in Burmese cats bred in America, which exhibit low T2D incidence. The original SNPs were confirmed in this cohort as associated with the T2D phenotype, although no novel coding SNPs in any of the seven candidate genes showed association with T2D. The identification of genetic markers associated with T2D susceptibility in ABB cats will enable preventative health strategies and guide breeding programs to reduce the prevalence of T2D in these cats.

From allozymes in 1983 to whole genomes in 2015, genetic studies of the cheetah have been extensive. In this chapter we provide an overview of the available literature. Overall, patterns of genetic variation provided evidence of low variability and suggest this loss occurred thousands of years ago. Differences between published subspecies were supported genetically. At a local scale, populations were generally considered panmictic with minor genetic structure. Although cheetahs have persisted despite low genetic variability, important questions arise from these findings: Does the cheetah have the ability to adapt to and evolve with future changes in environmental and infectious pressure? How would cheetahs cope with further loss of genetic diversity? Connectivity in the wild should be maintained via prevention of habitat loss, while management of small isolated populations may require reestablishing gene flow. Genetics could assist captive-breeding decisions and provide forensic evidence as to the geographical origin of illegally traded animals.

Diversity within the major histocompatibility complex (MHC) reflects the immunological fitness of a population. MHC-linked microsatellite markers provide a simple and an inexpensive method for studying MHC diversity in large-scale studies. We have developed 6 MHC-linked microsatellite markers in the domestic cat and used these, in conjunction with 5 neutral microsatellites, to assess MHC diversity in domestic mixed breed (n = 129) and purebred Burmese (n = 61) cat populations in Australia. The MHC of outbred Australian cats is polymorphic (average allelic richness = 8.52), whereas the Burmese population has significantly lower MHC diversity (average allelic richness = 6.81; P < 0.01). The MHC-linked microsatellites along with MHC cloning and sequencing demonstrated moderate MHC diversity in cheetahs (n = 13) and extremely low diversity in Gir lions (n = 13). Our MHC-linked microsatellite markers have potential future use in diversity and disease studies in other populations and breeds of cats as well as in wild felid species.

Over 200 hereditary diseases have been identified and reported in the cat, several of which affect the eye, with homology to human hereditary disease. Compared with traditional murine models, the cat demonstrates more features in common with humans, including many anatomic and physiologic similarities, longer life span, increased size, and a genetically more heterogeneous background. The development of genomic resources in the cat has facilitated mapping and further characterization of feline models. During recent years, the wealth of knowledge in feline ophthalmology and neurophysiology has been extended to include new diseases of significant interest for comparative ophthalmology. This makes the cat an extremely valuable animal species to utilize for further research into disease processes affecting both cats and humans. This is especially true in the advancement and study of new treatment regimens and for extended therapeutic trials. Groups of feline eye diseases reviewed in the following are lysosomal storage disorders, congenital glaucoma, and neuroretinal degenerations. Each has important implications for human ophthalmic research.

High-resolution genetic and physical maps are invaluable tools for building accurate genome assemblies, and interpreting results of genome-wide association studies (GWAS). Previous genetic and physical maps anchored good quality draft assemblies of the domestic cat genome, enabling the discovery of numerous genes underlying hereditary disease and phenotypes of interest to the biomedical science and breeding communities. However, these maps lacked sufficient marker density to order thousands of shorter scaffolds in earlier assemblies, which instead relied heavily on comparative mapping with related species. A high-resolution map would aid in validating and ordering chromosome scaffolds from existing and new genome assemblies. Here, we describe a high-resolution genetic linkage map of the domestic cat genome based on genotyping 453 domestic cats from several multi-generational pedigrees on the Illumina 63K SNP array. The final maps include 58,055 SNP markers placed relative to 6637 markers with unique positions, distributed across all autosomes and the X chromosome. Our final sex-averaged maps span a total autosomal length of 4464 cM, the longest described linkage map for any mammal, confirming length estimates from a previous microsatellite-based map. The linkage map was used to order and orient the scaffolds from a substantially more contiguous domestic cat genome assembly (Felis catus v8.0), which incorporated ∼20 × coverage of Illumina fragment reads. The new genome assembly shows substantial improvements in contiguity, with a nearly fourfold increase in N50 scaffold size to 18 Mb. We use this map to report probable structural errors in previous maps and assemblies, and to describe features of the recombination landscape, including a massive (∼50 Mb) recombination desert (of virtually zero recombination) on the X chromosome that parallels a similar desert on the porcine X chromosome in both size and physical location.

The glaucomas are a group of diseases characterized by optic nerve damage that together represent a leading cause of blindness in the human population and in domestic animals. Here we report a mutation in LTBP2 that causes primary congenital glaucoma (PCG) in domestic cats. We identified a spontaneous form of PCG in cats and established a breeding colony segregating for PCG consistent with fully penetrant, autosomal recessive inheritance of the trait. Elevated intraocular pressure, globe enlargement and elongated ciliary processes were consistently observed in all affected cats by 8 weeks of age. Varying degrees of optic nerve damage resulted by 6 months of age. Although subtle lens zonular instability was a common feature in this cohort, pronounced ectopia lentis was identified in less than 10% of cats examined. Thus, glaucoma in this pedigree is attributed to histologically confirmed arrest in the early post-natal development of the aqueous humor outflow pathways in the anterior segment of the eyes of affected animals. Using a candidate gene approach, significant linkage was established on cat chromosome B3 (LOD 18.38, θ = 0.00) using tightly linked short tandem repeat (STR) loci to the candidate gene, LTBP2. A 4 base-pair insertion was identified in exon 8 of LTBP2 in affected individuals that generates a frame shift that completely alters the downstream open reading frame and eliminates functional domains. Thus, we describe the first spontaneous and highly penetrant non-rodent model of PCG identifying a valuable animal model for primary glaucoma that closely resembles the human disease, providing valuable insights into mechanisms underlying the disease and a valuable animal model for testing therapies.

What Kitty Shares with Kings Although long-studied, the underlying basis of mammalian coat patterns remains unclear. By studying a large number of cat species and varieties, Kaelin et al. (p. 1536 ) identified two genes, Taqpep and Edn3 , as critical factors in the development of feline pigment patterns. Mutations in Taqpep are responsible for the blotched tabby pattern in domestic cats and the unusual coat of wild king cheetahs. Gene expression patterns in cat and cheetah skin suggest that Edn3 is a likely regulator of felid hair color. The findings support a common model for coat and pigment pattern formation in domestic and wild cats.

The use of DNA markers to identify sources of crime scene evidence has revolutionized forensic science. The routine use of simple tandem repeat (STR) markers applied to genetic individualization of human samples has become the realization of the potential of samples of nonhuman origin. The identification of an individual pet or other animal may provide the critical piece of information in a criminal investigation and prosecution. With the majority of American households (55%) housing at least one cat or dog (73 million cats, 68 million dogs) (1), it is not unusual for animal specimens, particularly hair specimens, to be part of the physical evidence associated with a crime scene (2–7). A study on the transfer and persistence of animal hair has demonstrated that it is almost impossible to enter a house where a domestic animal lives without becoming a “carrier” of its hair (8). Hairs from a pet can be indicative of a perpetrator’s presence at a crime scene or provide evidence of a connection between victim and perpetrator. Since 1996, DNA extracted from cat hair and dog blood, hair, and saliva from crime scene evidence has contributed to physical evidence leading to conviction of defendants in Canada and numerous states (2,4,5,9,10). While the genetic individualization of animal specimens is in its infancy, a set of guidelines has been proposed for animal genetic identity testing (11). We have developed an STR typing system for genetic individualization of domestic cat (Felis catus) samples. Previously, we reported on the characterization of a panel of 22 tetranucleotide STR loci in 213 domestic cats representing 28 breeds, from which 11 loci were selected for a forensic typing panel (12). A multiplex amplification protocol was generated and optimized for the 11 loci (12,13) and a gender-identifying sequence tagged site (STS) from the cat Y-chromosome Sex determining Region-Y gene (SRY) (14). We report on a population genetic database for the multiplex, with which to compute composite match probabilities, that was generated from a panel of 1043 domestic cat samples representing 38 cat breeds recognized by the two largest domestic cat registries in the United States, the Cat Fanciers’ Association (CFA) and The International Cat Association (TICA).

Only in recent years have specific mutations been elucidated for feline hereditary retinal dystrophies. Molecular genetic characterization of feline diseases has so far been a slow process but with a full genome sequence for the cat recently completed and the development of a feline single nucleotide polymorphism chip, the characterization of feline monogenic disorders will be significantly simplified. This review summarizes current knowledge with regard to specific hereditary retinal dystrophies in cats and gives an overview of how cats can be used as models in translational research.

Mammalian coat patterns (e.g., spots, stripes) are hypothesized to play important roles in camouflage and other relevant processes, yet the genetic and developmental bases for these phenotypes are completely unknown. The domestic cat, with its diversity of coat patterns, is an excellent model organism to investigate these phenomena. We have established three independent pedigrees to map the four recognized pattern variants classically considered to be specified by a single locus, Tabby; in order of dominance, these are the unpatterned agouti form called “Abyssinian” or “ticked” (Ta), followed by Spotted (Ts), Mackerel (TM), and Blotched (tb). We demonstrate that at least three different loci control the coat markings of the domestic cat. One locus, responsible for the Abyssinian form (herein termed the Ticked locus), maps to an ∼3.8-Mb region on cat chromosome B1. A second locus controls the Tabby alleles TM and tb, and maps to an ∼5-Mb genomic region on cat chromosome A1. One or more additional loci act as modifiers and create a spotted coat by altering mackerel stripes. On the basis of our results and associated observations, we hypothesize that mammalian patterned coats are formed by two distinct processes: a spatially oriented developmental mechanism that lays down a species-specific pattern of skin cell differentiation and a pigmentation-oriented mechanism that uses information from the preestablished pattern to regulate the synthesis of melanin profiles.

Objective To characterize hereditary retinal degeneration in the Abyssinian cat (rdAc) in a recently established closed colony segregating for the rdAc mutation, and evaluate possible differences in the age of onset and progression of disease phenotype since the initial description of rdAc 25 years ago. The sample size of an earlier study was increased in order to determine the allele frequency in Abyssinian and Somali cats on a worldwide basis. Animals studied Twenty rdAc affected cats from the closed animal facility, 87 Abyssinian and Somali cats for study of genotype–phenotype concordance, and DNA from 131 Abyssinian and Somali cats from Scandinavia, the UK and Australia for evaluation of the rdAc allele frequency. Procedures DNA was extracted from blood and buccal swabs using commercially available kits, followed by genotyping. Ophthalmic examinations were performed in the USA and Sweden by two board-certified veterinary ophthalmologists. Results A greater variation in the age of onset and progression of the disease was observed compared to that previously described. An excellent correlation between genotype and phenotype was observed. A population genetic survey revealed that the rdAc allele is in moderate abundance in the Abyssinian breed in Europe and Australia. Surprisingly, homozygosity for the mutant allele was observed in a Siamese cat with ophthalmoscopic findings similar to those originally described for affected rdAc individuals. Conclusions Alertness to the potential of rdAc is needed on the part of the veterinary ophthalmology community, not only in Abyssinian and Somali cats but possibly also in other related cat breeds.

Our knowledge of cat family biology was recently expanded to include a genomics perspective with the completion of a draft whole genome sequence of an Abyssinian cat. The utility of the new genome information has been demonstrated by applications ranging from disease gene discovery and comparative genomics to species conservation. Patterns of genomic organization among cats and inbred domestic cat breeds have illuminated our view of domestication, revealing linkage disequilibrium tracks consequent of breed formation, defining chromosome exchanges that punctuated major lineages of mammals and suggesting ancestral continental migration events that led to 37 modern species of Felidae. We review these recent advances here. As the genome resources develop, the cat is poised to make a major contribution to many areas in genetics and biology.

The world's domestic cats carry patterns of sequence variation in their genome that reflect a history of domestication and breed development. A genetic assessment of 979 domestic cats and their wild progenitors—Felis silvestris silvestris (European wildcat), F. s. lybica (Near Eastern wildcat), F. s. ornata (central Asian wildcat), F. s. cafra (southern African wildcat), and F. s. bieti (Chinese desert cat)—indicated that each wild group represents a distinctive subspecies of Felis silvestris. Further analysis revealed that cats were domesticated in the Near East, probably coincident with agricultural village development in the Fertile Crescent. Domestic cats derive from at least five founders from across this region, whose descendants were transported across the world by human assistance.

The historical literature suggests that in Australia, the domestic cat (Felis catus) had a European origin [~200 years before present (ybp)], but it is unclear if cats arrived from across the Asian land bridge contemporaneously with the dingo (4000 ybp), or perhaps immigrated ~40000 ybp in association with Aboriginal settlement from Asia. The origin of cats in Australia is important because the continent has a complex and ancient faunal assemblage that is dominated by endemic rodents and marsupials and lacks the large placental carnivores found on other large continents. Cats are now ubiquitous across the entire Australian continent and have been implicit in the range contraction or extinction of its small to medium sized (

The leading genetic cause of infant mortality is spinal muscular atrophy (SMA), a clinically and genetically heterogeneous group of disorders. Previously we described a domestic cat model of autosomal recessive, juvenile-onset SMA similar to human SMA type III. Here we report results of a whole-genome scan for linkage in the feline SMA pedigree using recently developed species-specific and comparative mapping resources. We identified a novel SMA gene candidate,LIX1, in an ~140-kb deletion on feline chromosome A1q in a region of conserved synteny to human chromosome 5q15. ThoughLIX1function is unknown, the predicted secondary structure is compatible with a role in RNA metabolism.LIX1expression is largely restricted to the central nervous system, primarily in spinal motor neurons, thus offering explanation of the tissue restriction of pathology in feline SMA. An exon sequence screen of 25 human SMA cases, not otherwise explicable by mutations at theSMN1locus, failed to identify comparableLIX1mutations. Nonetheless, aLIX1-associated etiology in feline SMA implicates a previously undetected mechanism of motor neuron maintenance and mandates consideration ofLIX1as a candidate gene in human SMA whenSMN1mutations are not found.

A forensic genotyping panel of 11 tetranucleotide STR loci from the domestic cat was characterized and evaluated for genetic individualization of cat tissues. We first examined 49 candidate STR loci and their frequency assessment in domestic cat populations. The STR loci (3–4 base pair repeat motifs), mapped in the cat genome relative to 579 coding loci and 255 STR loci, are well distributed across the 18 feline autosomes. All loci exhibit Mendelian inheritance in a multi-generation pedigree. Eleven loci that were unlinked and were highly heterozygous in cat breeds were selected for a forensic panel. Heterozygosity values obtained for the independent loci, ranged from 0.60–0.82, while the average cat breed heterozygosity obtained for the 11 locus panel was 0.71 (range of 0.57–0.83). A small sample set of outbred domestic cats displayed a heterozygosity of 0.86 for the 11 locus panel. The power of discrimination of the panel is moderate to high in the cat breeds examined, with an average Pm of 3.7E-06. The panel shows good potential for genetic individualization within outbred domestic cats with a Pm of 5.31E-08. A multiplex protocol, designed for the co-amplification of the 11 loci and a gender-identifying locus, is species specific and robust, generating a product profile with as little as 0.125 nanograms of genomic DNA.

The Dominant White locus (W) in the domestic cat demonstrates pleiotropic effects exhibiting complete penetrance for absence of coat pigmentation and incomplete penetrance for deafness and iris hypopigmentation. We performed linkage analysis using a pedigree segregating White to identify KIT (Chr. B1) as the feline W locus. Segregation and sequence analysis of the KIT gene in two pedigrees (P1 and P2) revealed the remarkable retrotransposition and evolution of a feline endogenous retrovirus (FERV1) as responsible for two distinct phenotypes of the W locus, Dominant White, and white spotting. A full-length (7125 bp) FERV1 element is associated with white spotting, whereas a FERV1 long terminal repeat (LTR) is associated with all Dominant White individuals. For purposes of statistical analysis, the alternatives of wild-type sequence, FERV1 element, and LTR-only define a triallelic marker. Taking into account pedigree relationships, deafness is genetically linked and associated with this marker; estimated P values for association are in the range of 0.007 to 0.10. The retrotransposition interrupts a DNAase I hypersensitive site in KIT intron 1 that is highly conserved across mammals and was previously demonstrated to regulate temporal and tissue-specific expression of KIT in murine hematopoietic and melanocytic cells. A large-population genetic survey of cats (n = 270), representing 30 cat breeds, supports our findings and demonstrates statistical significance of the FERV1 LTR and full-length element with Dominant White/blue iris (P < 0.0001) and white spotting (P < 0.0001), respectively.

The giant panda ( Ailuropoda melanoleuca ) is among the more familiar symbols of species conser- vation. The protection of giant panda populations has been aided recently by the establishment of more and better-managed reserves in existing panda habitat located in six mountain ranges in western China. These re- maining populations are becoming increasingly isolated from one another, however, leading to the concern that historic patterns of gene flow will be disrupted and that reduced population sizes will lead to diminished genetic variability. We analyzed four categories of molecular genetic markers (mtDNA restriction-fragment- length polymorphisms (RFLP), mtDNA control region sequences, nuclear multilocus DNA fingerprints, and mi- crosatellite size variation) in giant pandas from three mountain populations (Qionglai, Minshan, and Qin- ling) to assess current levels of genetic diversity and to detect evidence of historic population subdivisions. The three populations had moderate levels of genetic diversity compared with similarly studied carnivores for all four gene measures, with a slight but consistent reduction in variability apparent in the smaller Qinling population. That population also showed significant differentiation consistent with its isolation since historic times. From a strictly genetic perspective, the giant panda species and the three populations look promising insofar as they have retained a large amount of genetic diversity in each population, although evidence of re- cent population reduction—likely from habitat loss—is apparent. Ecological management to increase habi- tat, population expansion, and gene flow would seem an effective strategy to stabilize the decline of this en- dangered species.

The comparative method remains one of the most powerful approaches in the biological sciences (Harvey and Pagel 1991). The field of comparative genomics represents an invaluable extension of the Human Genome initiative, whereby genomic structure and function are interpreted in the evolutionary context of more manipulatable related animal species (O'Brien et al. 1997b,1999). Comparative gene maps serve two primary functions. First, they allow transfer of information from the gene-rich species maps of human and mouse to moderate-resolution gene maps of animal species that serve as models for biomedical, economical, and agricultural traits. Second, they provide a basis for understanding the rates and patterns of genome evolution (O'Brien et al. 1993; Andersson et al. 1996; VandeBerg and Graves 1998; O'Brien et al. 1999). Both of these endeavors require moderately dense comparative gene maps demarcating boundaries of conserved synteny among representative species. At present, three species, human, mouse, and rat, have gene-dense, ordered Type I maps for every chromosome (Copeland et al. 1993; Dietrich et al. 1997; Deloukas et al. 1998; Watanabe et al. 1999), producing a strong framework for comparing ordered gene maps of other mammalian species. However, the rodent genome comparisons appear to be rearranged two- to threefold more extensively than genomes from most other compared mammalian species genomes that display a highly conserved pattern of genome-wide conservation among mammalian Orders (Graves 1996; Carver and Stubbs 1997; O'Brien et al. 1999). Other than rodents, most whole genome comparisons to humans have been limited to partial maps on the basis of unordered somatic cell hybrid or Zoo-FISH (chromosome painting)-based synteny, which do not reveal intrachromosomal inversions that have occurred during the evolution of these species (Chowdhary et al. 1998; Ferguson-Smith et al. 1998; Wienberg and Stanyon 1998; Finelli et al. 1999). It is becoming clear that a complete view of the extent and character of genome rearrangements that have occurred during species evolution must include precise orders of compared Type I-coding genes on each chromosome in different mammalian genomes (O'Brien 1991; Copeland et al. 1993; Womack and Kata 1995; Schibler et al. 1997; Yang and Womack 1998; Murphy et al. 1999a,b; Watanabe et al. 1999). The domestic cat has long played a role in comparative medicine beginning with comparative anatomy, physiology, and biochemistry, extending to the present, in which the species provides numerous models for human heritable and infectious disease (http://www.angis.org.au/Databases/BIRX/omia/) including lysosomal storage diseases (Yogalingam et al. 1996; Berg et al. 1997; Fyfe et al. 1999), polycystic kidney disease (Biller 1996), and AIDS (FIV-feline immunodeficiency virus; Willett et al. 1997). The domestic cat also provides a valuable reference for comparative genome analysis, displaying remarkable syntenic conservation relative to the human genome (O'Brien et al. 1997a,b, 1999; Rettenberger et al. 1995; Wienberg et al. 1997). As for most published comparative mapping efforts in other species, these cat–human synteny observations are based on somatic cell-hybrid mapping, FISH, and Zoo-FISH results, but cannot reveal underlying subchromosomal conservation or rearrangement relative to other species. Whole-genome radiation hybrid (RH) mapping has become a powerful and expedient method for gene ordering in vertebrate species by largely eliminating the need to identify within-species polymorphism (Cox et al. 1990; Walter et al. 1994). To date, this approach has been used to generate genome-spanning maps for the human, rat, and mouse genomes (Stewart et al. 1997; Deloukas et al. 1998; Steen et al. 1999; Van Etten et al. 1999; Watanabe et al. 1999), although maps are beginning to be developed for other mammalian species (Radiation Hybrid Database: www.linkage.rockefeller.edu/tara/rhmap). Using the feline RH panel (Murphy et al. 1999a), we present an RH map for the domestic cat that integrates 424 Type I (coding) loci with 176 selected Type II microsatellite loci ordered previously in an interspecies backcross-generated linkage map (Menotti-Raymond et al. 1999). This map represents the first ordered, full-genome parallel RH comparison to the human genome for a non-rodent species. The integrated 600 marker map has a mean density of 5 cM/26 cR5000 now useful for comparative candidate positional clone analysis of the >200 described feline hereditary phenotypes. A comparison of human to feline conserved segments revealed 100 ordered segments across both species genomes, more than three times the number revealed by previous comparative genomic methodologies (O'Brien et al. 1997a,b; Rettenberger et al. 1995; Wienberg et al. 1997). These data demonstrate the increased comparative precision required for accurate evolutionary inference and the achievement of this precision by moderately dense RH maps of Type I-coding gene markers.

[Figure][1] Traces of evolutionary history appear in functional morphology and DNA sequences of living and extinct species. These remnants of the past can lead to insights into the relationships among extant groups of animals, the forces driving evolution, and the utility of animal models for studying human disease. We present below one evolutionary interpretation of the still-disputed hierarchy of surviving placental mammalian orders (excluding monotremes and marsupials), a synthesis of accumulated molecular and morphological inferences. The time scale is derived largely from molecular data; indicated fossil remains are much younger, raising controversies around the precise age of mammal ancestors. The tips of the phylogenetic tree depict genomes for 21 representative species from 11 orders, which are labeled on the vertical lines. These examples show genome-wide homology alignments assessed by comparative gene mapping or direct visualization of chromosome painting, in which fluorescently labeled, individual chromosomes are hybridized to chromosomes from distantly related species. Species were selected to maximize the number of orders represented and to illustrate patterns of genome conservation. The 24 human chromosomes are distinguished by 24 colors, and regions of human chromosome homology in other species are indicated by color and human chromosome number (for example, cat chromosome A1 contains gene stretches homologous to human chromosomes 5 and 13). Selected gene homologs in each species represent a subset of extensive comparative gene mapping data; thus genes listed may not reflect the extreme borders of the conserved segments. Gene orders have been determined in human, cat, pig, sheep, rat, mouse, and zebrafish; in other species the genes are listed in the order in which they appear in humans. The zebrafish map, including 25 linkage groups not yet assigned to the 25 chromosomes, is presented to illustrate the array of gene segments conserved for 450 million years. Further information and discussion of dissenting views of mammalian evolution can be found in this issue of Science and the associated web site at ![Figure][1] CREDITS Science Coordinators: Barbara R. Jasny and Pamela J. Hines Authors: Phylogenetic tree Stephen J. O'Brien, National Cancer Institute, Frederick, MD, USA; John F. Eisenberg, University of Florida, Gainesville, FL, USA; Michael Miyamoto, University of Florida, Gainesville, FL, USA; S. Blair Hedges, Pennsylvania State University, University Park, PA, USA; Sudhir Kumar, Arizona State University, Tempe, AZ, USA; Don E. Wilson, Smithsonian Institution, Washington, DC, USA. Genomic maps Stephen J. O'Brien, Marilyn Menotti-Raymond, William J. Murphy, William G. Nash, Leslie A. Lyons, Joan C. Menninger, Roscoe Stanyon, Johannes Wienberg, Neal G. Copeland, Nancy A. Jenkins, National Cancer Institute, Frederick, MD, USA; Joel Gellin, Martine Yerle, Institut National de la Recherche Agronomique, Castanet-Tolosan, France; Leif Andersson, Swedish University of Agricultural Sciences, Uppsala, Sweden; James Womack, Texas A & M University, College Station, TX, USA; Thomas Broad, AgResearch, Invermay, Mosgiel, New Zealand; John Postlethwait, University of Oregon, Eugene, OR, USA; Oleg Serov, Institute of Cytology and Genetics, Siberian Branch of the Academy of Sciences of Russia, Novosibirsk, Russia; Ernie Bailey, University of Kentucky, Lexington, KY, USA; Michael R. James, Wellcome Trust Centre for Human Genetics, Headington, UK; Takeshi K. Watanabe, Otsuka GEN Research Institute, Tokushima, Japan; Matthew J. Wakefield, Jennifer Marshall Graves, La Trobe University, Melbourne, Australia. Design and Art Direction: C. Faber Smith Graphics: Nathalie Cary Illustrations: Katharine Sutliff Production Assistance: Holly Bishop, Leslie Blizard, Inja Lin, Darcel Pugh Proofreading: Harry Jach [1]: pending:yes

Dense genetic maps of human, mouse, and rat genomes that are based on coding genes and on microsatellite and single-nucleotide polymorphism markers have been complemented by precise gene homolog alignment with moderate-resolution maps of livestock, companion animals, and additional mammal species. Comparative genetic assessment expands the utility of these maps in gene discovery, in functional genomics, and in tracking the evolutionary forces that sculpted the genome organization of modern mammalian species.

U poglavlju se opisuje primjena analiza životinjske DNA k oja se primjenjuje u kriminalistickim slucajevima, utvrđivanju pedigrea životinja te u forenzickim slucajevima koji ukljucuju divlje životinje

A simple tandem repeat (STR) PCR-based typing system developed for the genetic individualization of domestic cat samples has been used to generate a population genetic database of domestic cat breeds. A panel of 10 tetranucleotide STR loci and a gender-identifying sequence tagged site (STS) were co-amplified in genomic DNA of 1043 individuals representing 38 cat breeds. The STR panel exhibits relatively high heterozygosity in cat breeds, with an average 10-locus heterozygosity of 0.71, which represents an average of 38 breed-specific heterozygosities for the 10-member panel. When the entire set of breed individuals was analyzed as a single population, a heterozygosity of 0.87 was observed. Heterozygosities obtained for the 10 loci range from 0.72 to 0.96. The power for genetic individualization of domestic cat samples of the multiplex is high, with a probability of match (p(m)) of 6.2E-14, using a conservative θ = 0.05.

Recent advances in human gene mapping technologies have led to increased emphasis in developing representative genetic maps for several species, particularly domestic plants and animals. These maps are being compiled with two distinct goals: to provide a resource for genetic analysis, and to help dissect the evolution of genome organization by comparing linkage relationships of homologous genes. We recently proposed a list of reference anchor loci suitable for comparative gene mapping in mammals and other vertebrate classes. The comparative anchors, which are Type I (coding gene) loci, provide a comparative framework, but the addition of Type II (microsatellite) loci is also necessary for mapping phenotypes in animal gene maps. Strategies for combining the advantages of Type I and Type II locus markers are presented with emphasis on application of these concepts to the construction of a gene map of the domestic cat.

Only in recent years have specific mutations been elucidated for feline hereditary retinal dystrophies. Molecular genetic characterization of feline diseases has so far been a slow process but with a full genome sequence for the cat recently completed and the development of a feline single nucleotide polymorphism chip, the characterization of feline monogenic disorders will be significantly simplified. This review summarizes current knowledge with regard to specific hereditary retinal dystrophies in cats and gives an overview of how cats can be used as models in translational research.

The cheetah is unusual among fields in exhibiting near genetic uniformity at a variety of loci previously screened to measure population genetic diversity. It has been hypothesized that a demographic crash or population bottleneck in the recent history of the species is causal to the observed monomorphic profiles for nuclear coding loci. The timing of a bottleneck is difficult to assess, but certain aspects of the cheetah's natural history suggest it may have occurred near the end of the last ice age (late Pleistocene, approximately 10,000 years ago), when a remarkable extinction of large vertebrates occurred on several continents. To further define the timing of such a bottleneck, the character of genetic diversity for two rapidly evolving DNA sequences, mitochondrial DNA and hypervariable minisatellite loci, was examined. Moderate levels of genetic diversity were observed for both of these indices in surveys of two cheetah subspecies, one from South Africa and one from East Africa. Back calculation from the extent of accumulation of DNA diversity based on observed mutation rates for VNTR (variable number of tandem repeats) loci and mitochondrial DNA supports a hypothesis of an ancient Pleistocene bottleneck that rendered the cheetah depauperate in genetic variation for nuclear coding loci but would allow sufficient time for partial reconstitution of more rapidly evolving genomic DNA segments.

The recent discovery of a mutational variant in the CEP290 gene (CEP290: IVS50 + 9T>G), conferring recessive retinal degeneration in Abyssinian and Somali (long-haired Abyssinian) cats (rdAc) prompted a survey among 41 cat breeds (846 individuals) to assess the incidence, frequency and clinical consequence of rdAc. The rdAc allele displayed widespread distribution, observed in 16/43 (37%) breeds, exhibiting a high allele frequency (~33%) in North American and European Siamese populations. Clinical evaluations demonstrated high concordance between rdAc pathology and the CEP290 (IVS50 + 9T>G) homozygous genotype (P = 1.1E–6), with clinical disease similar to affected Abyssinians/Somalis. This retinal degeneration has not been reported in breeds other than the Abyssinian/Somali and poses a significant health risk particularly in the Siamese breed group. Alertness of the veterinary community and the present availability of commercial diagnostic testing could synergistically enable breeders to reduce the incidence of rdAc blindness in pure-bred cat populations.

The SILVER locus has been mapped in the domestic cat, identifying a unique genomic location distinct from that of any known reported gene associated with silver or hypopigmentation in mammals. A demonstrated lack of linkage to SILV, the strong candidate gene for silver, led to the initiation of a genome scan utilizing 2 pedigrees segregating for silver coat color. Linkage mapping defined a genomic region for SILVER as a 3.3-Mb region, (95.87–99.21 Mb) on chromosome D2, (peak logarithm of the odds = 10.5, θ = 0), which displays conserved synteny to a genomic interval between 118.58 and 121.85 Mb on chromosome 10 in the human genome. In the domestic cat, mutations at the SILVER locus suppress the development of pigment in the hair, but in contrast to other mammalian silver variants, there is an apparently greater influence on the production of pheomelanin than eumelanin pigment. The mapping of a novel locus for SILVER offers much promise in identifying a gene that may help elucidate aspects of pheomelanogenesis, a pathway that has been very elusive, and illustrates the promise of the cat genome project in increasing our understanding of basic biological processes of general relevance for mammals.

A comprehensive genetic linkage map of the domestic cat X chromosome was generated with the goal of localizing the genomic position of the classic X-linked orange (O) locus. Microsatellite markers with an average spacing of 3 Mb were selected from sequence traces of the cat 1.9× whole genome sequence (WGS), including the pseudoautosomal region 1 (PAR1). Extreme variation in recombination rates (centimorgans per megabase) was observed along the X chromosome, ranging from a virtual absence of recombination events in a region estimated to be >30 Mb to recombination frequencies of 15.7 cM/Mb in a segment estimated to be

We report on the completion of an autosomal genetic linkage (GL) map of the domestic cat (Felis silvestris catus). Unlike two previous linkage maps of the cat constructed with a hybrid pedigree between the domestic cat and the Asian leopard cat, this map was generated entirely with domestic cats, using a large multi-generational pedigree (n = 256) maintained by the Nestle Purina PetCare Company. Four hundred eighty-three simple tandem repeat (STR) loci have been assigned to linkage groups on the cat's 18 autosomes. A single linkage group spans each autosome. The length of the cat map, estimated at 4370 cM, is long relative to most reported mammalian maps. A high degree of concordance in marker order was observed between the third-generation map and the 1.5 Mb-resolution radiation hybrid (RH) map of the cat. Using the cat 1.9 × whole-genome sequence, we identified map coordinates for 85% of the loci in the cat assembly, with high concordance observed in marker order between the linkage map and the cat sequence assembly. The present version represents a marked improvement over previous cat linkage maps as it (i) nearly doubles the number of markers that were present in the second-generation linkage map in the cat, (ii) provides a linkage map generated in a domestic cat pedigree which will more accurately reflect recombination distances than previous maps generated in a hybrid pedigree, and (iii) provides single linkage groups spanning each autosome. Marker order was largely consistent between this and the previous maps, though the use of a hybrid pedigree in the earlier versions appears to have contributed to some suppression of recombination. The improved linkage map will provide an added resource for the mapping of phenotypic variation in the domestic cat and the use of this species as a model system for biological research.

The domestic cat, currently the most frequent of companion animals, has enjoyed a medical surveillance, as a nonprimate species, second only to the dog. With over 200 hereditary disease pathologies reported in the cat, the clinical and physiological study of these feline hereditary diseases provides a strong comparative medicine opportunity for prevention, diagnostics, and treatment studies in a laboratory setting. Causal mutations have been characterized in 19 felid genes, with the largest representation from lysosomal storage enzyme disorders. Corrective therapeutic strategies for several disorders have been proposed and examined in the cat, including enzyme replacement, heterologous bone marrow transplantation, and substrate reduction therapy. Genomics tools developed in the cat, including the recent completion of the 2-fold whole genome sequence of the cat and genome browser, radiation hybrid map of 1793 integrated coding and microsatellite loci, a 5-cM genetic linkage map, arrayed BAC libraries, and flow sorted chromosomes, are providing resources that are being utilized in mapping and characterization of genes of interest. A recent report of the mapping and characterization of a novel causative gene for feline spinal muscular atrophy marked the first identification of a disease gene purely from positional reasoning. With the development of genomic resources in the cat and the application of complementary comparative tools developed in other species, the domestic cat is emerging as a promising resource of phenotypically defined genetic variation of biomedical significance. Additionally, the cat has provided several useful models for infectious disease. These include feline leukemia and feline sarcoma virus, feline coronavirus, and Type C retroviruses that interact with cellular oncogenes to induce leukemia, lymphoma, and sarcoma.

The genome sequence (1.9-fold coverage) of an inbred Abyssinian domestic cat was assembled, mapped, and annotated with a comparative approach that involved cross-reference to annotated genome assemblies of six mammals (human, chimpanzee, mouse, rat, dog, and cow). The results resolved chromosomal positions for 663,480 contigs, 20,285 putative feline gene orthologs, and 133,499 conserved sequence blocks (CSBs). Additional annotated features include repetitive elements, endogenous retroviral sequences, nuclear mitochondrial (numt) sequences, micro-RNAs, and evolutionary breakpoints that suggest historic balancing of translocation and inversion incidences in distinct mammalian lineages. Large numbers of single nucleotide polymorphisms (SNPs), deletion insertion polymorphisms (DIPs), and short tandem repeats (STRs), suitable for linkage or association studies were characterized in the context of long stretches of chromosome homozygosity. In spite of the light coverage capturing ∼65% of euchromatin sequence from the cat genome, these comparative insights shed new light on the tempo and mode of gene/genome evolution in mammals, promise several research applications for the cat, and also illustrate that a comparative approach using more deeply covered mammals provides an informative, preliminary annotation of a light (1.9-fold) coverage mammal genome sequence.

To conduct developmental validation studies on a polymerase chain reaction (PCR) based short tandem repeat (STR) multiplex typing system, developed for the purpose of genetic individualization and parentage testing in domestic cat samples.To evaluate reproducibility of the typing system, the multiplex was amplified using DNA extracted from hair, blood, and buccal samples obtained from the same individual (n=13). Additional studies were performed to evaluate the system's species' specificity, using 26 North American mammalian species and two prokaryotes Sacchromyces and Escherichia coli, sensitivity, and ability to identify DNA mixtures. Patterns of Mendelian inheritance and mutation rates for the 11 loci were directly examined in a large multi-generation domestic cat pedigree (n=263).Our studies confirm that the multiplex system was species-specific for feline DNA and amplified robustly with as little as 125 picograms of genomic template DNA, demonstrating good product balance. The multiplex generated all components of a two DNA mixture when the minor component was one tenth of the major component at a threshold of 50 relative fluorescence units. The multiplex was reproducible in multiple tissue types of the same individual. Mutation rates for the 11 STR were within the range of sex averaged rates observed for Combined DNA Index System (CODIS) loci.The cat STR multiplex typing system is a robust and reliable tool for the use of forensic DNA analysis of domestic cat samples.

Y chromosome haplotyping based on microsatellites and single nucleotide polymorphisms (SNPs) has proved to be a powerful tool for population genetic studies of humans. However, the promise of the approach is hampered in the majority of nonhuman mammals by the lack of Y-specific polymorphic markers. We were able to identify new male-specific polymorphisms in the domestic cat Felis catus and 6 additional Felidae species with a combination of molecular genetic and cytogenetic approaches including 1) identifying domestic cat male-specific microsatellites from markers generated from a male cat microsatellite-enriched genomic library, a flow-sorted Y cosmid library, or a Y-specific cat bacteria artificial chromosome (BAC) clone, (2) constructing microsatellite-enriched libraries from flow-sorted Y chromosomes isolated directly from focal wildcat species, and (3) screening Y chromosome conserved anchored tagged sequences primers in Felidae species. Forty-one male-specific microsatellites were identified, but only 6 were single-copy loci, consistent with the repetitive nature of the Y chromosome. Nucleotide diversity (pi) of Y-linked intron sequences (2.1 kbp) was in the range of 0 (tiger) to 9.95 x 10(-4) (marbled cat), and the number of SNPs ranged from none in the tiger to 7 in the Asian leopard cat. The Y haplotyping system described here, consisting of 4 introns (SMCY3, SMCY7, UTY11, and DBY7) and 1 polymorphic microsatellite (SMCY-STR), represents the first available markers for tracking intraspecific male lineage polymorphisms in Felidae species and promises to provide significant insights to evolutionary and population genetic studies of the species.

Genetic variation in cat breeds was assessed utilizing a panel of short tandem repeat (STR) loci genotyped in 38 cat breeds and 284 single-nucleotide polymorphisms (SNPs) genotyped in 24 breeds. Population structure in cat breeds generally reflects their recent ancestry and absence of strong breed barriers between some breeds. There is a wide range in the robustness of population definition, from breeds demonstrating high definition to breeds with as little as a third of their genetic variation partitioning into a single population. Utilizing the STRUCTURE algorithm, there was no clear demarcation of the number of population subdivisions; 16 breeds could not be resolved into independent populations, the consequence of outcrossing in established breeds to recently developed breeds with common ancestry. These 16 breeds were divided into 6 populations. Ninety-six percent of cats in a sample set of 1040 were correctly assigned to their classified breed or breed group/population. Average breed STR heterozygosities ranged from moderate (0.53; Havana, Korat) to high (0.85; Norwegian Forest Cat, Manx). Most of the variation in cat breeds was observed within a breed population (83.7%), versus 16.3% of the variation observed between populations. The hierarchical relationships of cat breeds is poorly defined as demonstrated by phylogenetic trees generated from both STR and SNP data, though phylogeographic grouping of breeds derived completely or in part from Southeast Asian ancestors was apparent.

To determine the genetic regulation of "hair length" in the domestic cat, a whole-genome scan was performed in a multigenerational pedigree in which the "long-haired" phenotype was segregating. The 2 markers that demonstrated the greatest linkage to the long-haired trait (log of the oddsor = 6) flanked an estimated 10-Mb region on cat chromosome B1 containing the Fibroblast Growth Factor 5 (FGF5) gene, a candidate gene implicated in regulating hair follicle growth cycle in other species. Sequence analyses of FGF5 in 26 cat breeds and 2 pedigrees of nonbreed cats revealed 4 separate mutations predicted to disrupt the biological activity of the FGF5 protein. Pedigree analyses demonstrated that different combinations of paired mutant FGF5 alleles segregated with the long-haired phenotype in an autosomal recessive manner. Association analyses of more than 380 genotyped breed and nonbreed cats were consistent with mutations in the FGF5 gene causing the long-haired phenotype in an autosomal recessive manner. In combination, these genomic approaches demonstrated that FGF5 is the major genetic determinant of hair length in the domestic cat.

Juvenile-onset spinal muscular atrophy was observed in an extended family of purebred domestic cats as a fully penetrant, simple autosomal recessive trait. Affected kittens exhibited tremor, proximal muscle weakness, and muscle atrophy beginning at ~4 mo of age. Apparent loss of function was rapid initially but progressed slowly after 7-8 mo of age, and variably disabled cats lived for at least 8 y. Electromyography and microscopic examination of muscle and nerve biopsies were consistent with denervation atrophy as a result of a central lesion. There was astrogliosis and dramatic loss of motor neurons in ventral but not dorsal horn gray matter of spinal cord and loss of axons in ventral horn nerve roots. These phenotypic findings were similar to mild forms (type III) of spinal muscular atrophy in humans caused by survival of motor neuron mutations, but molecular analysis excluded feline survival of motor neuron as the disease gene in this family. A breeding colony has been established for further investigation of this naturally occurring large-animal model of inherited motor neuron disease.

A quantitative polymerase chain reaction (PCR) assay has been developed for the quantification of genomic DNA extracted from domestic cat samples. The assay, which targets highly repetitive genomic short interspersed nuclear elements (SINE), can be performed rapidly and is highly sensitive, detecting as little as 10 fg of feline genomic DNA. The assay was linear over a 10(6) dilution range. We have recently developed a short tandem repeat (STR) multiplex panel for forensic analysis of feline specimens. The SINE assay is an integral part of the forensic typing system. The sensitivity of the assay will enable forensic examiners to determine the likelihood of success of genotyping sample extracts with the STR panel without sacrificing valuable DNA necessary to perform genotyping of samples.

Effective utilization of the domestic cat as an animal model for hereditary and infectious disease requires the development and implementation of high quality gene maps incorporating microsatellites and conserved coding gene markers. Previous feline linkage and radiation hybrid maps have lacked sufficient microsatellite coverage on all chromosomes to make effective use of full genome scans. Here we report the isolation and genomic mapping of 304 novel polymorphic repeat loci in the feline genome. The new loci were mapped in the domestic cat radiation hybrid panel using an automated fluorescent Taq-Man based assay. The addition of these 304 microsatellites brings the total number of microsatellites mapped in the feline genome to 580, and the total number of loci placed onto the RH map to 1,126. Microsatellites now span every autosome with an average spacing of roughly one polymorphic STR every five centimorgans, and full genome coverage of one marker every 2.7 megabases. These loci now provide a useful tool for undertaking full-genome scans to identify genes associated with phenotypes of interest, such as those relating to hereditary disease, coat color, patterning and morphology. These resources can also be extended to the remaining 36 species of the cat family for population genetic and evolutionary genomic analyses.

Melanistic coat coloration occurs as a common polymorphism in 11 of 37 felid species and reaches high population frequency in some cases but never achieves complete fixation [1–3]. To investigate the genetic basis, adaptive significance, and evolutionary history of melanistic variants in the Felidae, we mapped, cloned, and sequenced the cat homologs of two putative candidate genes for melanism (ASIP [agouti] and MC1R) and identified three independent deletions associated with dark coloration in three different felid species. Association and transmission analyses revealed that a 2 bp deletion in the ASIP gene specifies black coloration in domestic cats, and two different “in-frame” deletions in the MC1R gene are implicated in melanism in jaguars and jaguarundis. Melanistic individuals from five other felid species did not carry any of these mutations, implying that there are at least four independent genetic origins for melanism in the cat family. The inferred multiple origins and independent historical elevation in population frequency of felid melanistic mutations suggest the occurrence of adaptive evolution of this visible phenotype in a group of related free-ranging species.

▪ Abstract The compilation of a dense gene map and eventually a whole genome sequence (WGS) of the domestic cat holds considerable value for human genome annotation, for veterinary medicine, and for insight into the evolution of genome organization among mammals. Human association and veterinary studies of the cat, its domestic breeds, and its charismatic wild relatives of the family Felidae have rendered the species a powerful model for human hereditary diseases, for infectious disease agents, for adaptive evolutionary divergence, for conservation genetics, and for forensic applications. Here we review the advantages, rationale, and present strategy of a feline genome project, and we describe the disease models, comparative genomics, and biological applications posed by the full resolution of the cat's genome.